WO2016044400A1 - Micelles à noyau de coacervat complexe à copolymère bloc pour la catalyse enzymatique dans un solvant organique - Google Patents

Micelles à noyau de coacervat complexe à copolymère bloc pour la catalyse enzymatique dans un solvant organique Download PDF

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WO2016044400A1
WO2016044400A1 PCT/US2015/050387 US2015050387W WO2016044400A1 WO 2016044400 A1 WO2016044400 A1 WO 2016044400A1 US 2015050387 W US2015050387 W US 2015050387W WO 2016044400 A1 WO2016044400 A1 WO 2016044400A1
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nanostructure
enzyme
block copolymer
acid
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Bradley David OLSEN
Carolyn Elaine MILLS
Xuehui Dong
Aliie Caitlin OBERMEYER
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Massachusetts Institute Of Technology
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    • A62D3/00—Processes for making harmful chemical substances harmless or less harmful, by effecting a chemical change in the substances
    • A62D3/02—Processes for making harmful chemical substances harmless or less harmful, by effecting a chemical change in the substances by biological methods, i.e. processes using enzymes or microorganisms
    • A—HUMAN NECESSITIES
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    • A61K9/1075—Microemulsions or submicron emulsions; Preconcentrates or solids thereof; Micelles, e.g. made of phospholipids or block copolymers
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    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J13/00—Colloid chemistry, e.g. the production of colloidal materials or their solutions, not otherwise provided for; Making microcapsules or microballoons
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    • B—PERFORMING OPERATIONS; TRANSPORTING
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    • B01J31/00—Catalysts comprising hydrides, coordination complexes or organic compounds
    • B01J31/003—Catalysts comprising hydrides, coordination complexes or organic compounds containing enzymes
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J31/00—Catalysts comprising hydrides, coordination complexes or organic compounds
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    • C11D—DETERGENT COMPOSITIONS; USE OF SINGLE SUBSTANCES AS DETERGENTS; SOAP OR SOAP-MAKING; RESIN SOAPS; RECOVERY OF GLYCEROL
    • C11D3/00—Other compounding ingredients of detergent compositions covered in group C11D1/00
    • C11D3/16—Organic compounds
    • C11D3/37—Polymers
    • C11D3/3746—Macromolecular compounds obtained by reactions only involving carbon-to-carbon unsaturated bonds
    • C11D3/378—(Co)polymerised monomers containing sulfur, e.g. sulfonate
    • C—CHEMISTRY; METALLURGY
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    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
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    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/24—Hydrolases (3) acting on glycosyl compounds (3.2)
    • C12N9/2402—Hydrolases (3) acting on glycosyl compounds (3.2) hydrolysing O- and S- glycosyl compounds (3.2.1)
    • C12N9/2405—Glucanases
    • C12N9/2408—Glucanases acting on alpha -1,4-glucosidic bonds
    • C12N9/2411—Amylases
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    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/24—Hydrolases (3) acting on glycosyl compounds (3.2)
    • C12N9/2402—Hydrolases (3) acting on glycosyl compounds (3.2) hydrolysing O- and S- glycosyl compounds (3.2.1)
    • C12N9/2405—Glucanases
    • C12N9/2408—Glucanases acting on alpha -1,4-glucosidic bonds
    • C12N9/2411—Amylases
    • C12N9/2414—Alpha-amylase (3.2.1.1.)
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    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/48—Hydrolases (3) acting on peptide bonds (3.4)
    • C12N9/50—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25)
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    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/48—Hydrolases (3) acting on peptide bonds (3.4)
    • C12N9/50—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25)
    • C12N9/64—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25) derived from animal tissue
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    • C12Y—ENZYMES
    • C12Y301/00—Hydrolases acting on ester bonds (3.1)
    • C12Y301/08—Phosphoric triester hydrolases (3.1.8)
    • C12Y301/08001—Aryldialkylphosphatase (3.1.8.1), i.e. paraoxonase
    • A—HUMAN NECESSITIES
    • A62—LIFE-SAVING; FIRE-FIGHTING
    • A62D—CHEMICAL MEANS FOR EXTINGUISHING FIRES OR FOR COMBATING OR PROTECTING AGAINST HARMFUL CHEMICAL AGENTS; CHEMICAL MATERIALS FOR USE IN BREATHING APPARATUS
    • A62D2101/00—Harmful chemical substances made harmless, or less harmful, by effecting chemical change
    • A62D2101/02—Chemical warfare substances, e.g. cholinesterase inhibitors
    • A—HUMAN NECESSITIES
    • A62—LIFE-SAVING; FIRE-FIGHTING
    • A62D—CHEMICAL MEANS FOR EXTINGUISHING FIRES OR FOR COMBATING OR PROTECTING AGAINST HARMFUL CHEMICAL AGENTS; CHEMICAL MATERIALS FOR USE IN BREATHING APPARATUS
    • A62D2101/00—Harmful chemical substances made harmless, or less harmful, by effecting chemical change
    • A62D2101/20—Organic substances
    • A62D2101/26—Organic substances containing nitrogen or phosphorus

Definitions

  • Enzymes or enzyme clusters can be isolated in a variety of nanostructures, such as viral capsids, reverse micelles, and polymersomes. Studying such encapsulated enzymes has shed light on enzymatic behavior in the absence of bulk aqueous solution. Nanostructures may also be used to solubilize directly and efficiently protein clusters into organic solvents containing small quantities of surfactant and trace amounts of water. As a result of this stabilization, enzymes may also exhibit increased enzyme activity relative to extracted enzyme activity. This approach is appealing for bioreactor fabrication; of particular interest is the fabrication of bioreactors capable of efficiently and effectively sequestering and eliminating dangerous chemicals, such as nerve agents. However, there are critical issues regarding the regulation of solute transport through membranes of the nanostructure, enzyme loading without denaturation, and physiological stability.
  • bioreactors capable of efficiently and effectively sequestering and eliminating dangerous chemicals, such as nerve agents.
  • the invention relates to a nanostructure, comprising, consisting essentially of, or consisting of:
  • the invention relates to any one of the nanostructures described herein, wherein the block copolymer comprises a plurality of first repeat units, and a plurality of second repeat units
  • the first repeat unit is ,
  • R is H, alkyl, halo, hydroxy, amino, nitro, or
  • Y is alkyl
  • ⁇ is an anion
  • R is H, alkyl, halo, hydroxy, amino, nitro, or cyano
  • p 2-20, inclusive.
  • the invention relates to any one of the nanostructures described herein, wherein the enzyme or modified enzyme is an organophosphate hydrolase or a modified organophosphate hydrolase.
  • the invention relates to any one of the nanostructures described herein, wherein the enzyme or modified enzyme is an organophosphate acid anhydrolase or a modified organophosphate acid anhydrolase.
  • the invention relates to a composition, comprising, consisting essentially of, or consisting of:
  • the invention relates to a method of hydrolyzing an organophosphorous compound, comprising contacting the organophosphorous compound with an effective amount of any of the nanostructures or compositions described herein. In certain embodiments, the invention relates to a method of decontaminating an area or a device contaminated with an organophosphorous compound, comprising contacting the area or the device with an effective amount of any of the nanostructures or compositions described herein.
  • Figure 1A depicts the hydrolysis of sarin by organophosphate hydrolase (OPH).
  • OHP organophosphate hydrolase
  • Figure IB depicts the hydrolysis of VX by organophosphate acid anhydrolase (OPAA).
  • Figure 2 is a schematic representation of a complex coacervate core micelle acting as a nanoreactor.
  • Figure 3 depicts an exemplary synthesis of POEGMA-b-quaternized P4VP.
  • FIG 4 is a schematic representation of a general method of forming complex coacervate core micelles (C3Ms) with a block copolymer, such as POEGMA-b-qP4VP, and a protein, such as organophosphate hydrolase.
  • C3Ms complex coacervate core micelles
  • Figure 5 is a schematic representation of a general method of forming C3Ms with a block copolymer and a supercharged protein.
  • Figure 6A depicts a method of supercharging a protein comprising a lysine residue.
  • Figure 6B depicts the zeta potential of supercharged proteins as compared to unmodified proteins.
  • Figure 7A depicts two components, a supercharged protein and a quaternized P4VP homopolymer, used to investigate coacervation.
  • Figure 7B depicts the results of dynamic light scattering (DLS) studies at 600 nm of the components from Figure 7A in 50 mM phosphate buffer at pH 7.0, 8.0, and 9.0, as a function of weight fraction qP4VP.
  • DLS dynamic light scattering
  • Figure 9 depicts the results of DLS studies at 600 nm of the components from Figure 7A in 50 mM phosphate buffer at pH 8.0, as a function of weight fraction qP4VP. The 0.5 weight fraction was selected for DLS studies with the block copolymer.
  • Figure 10A depicts two components, a supercharged protein and a POEGMA-b- qP4VP block copolymer, used to investigate micelle formation.
  • Figure 11 is a schematic representation of a general method for forming C3Ms with a block copolymer, a protein, and a charged homopolymer.
  • Figure 12A depicts two components, a charged homopolymer and a quaternized P4VP homopolymer, used to investigate coacervation.
  • Figure 12B depicts the results of DLS studies at 600 nm of the components from Figure 12A in 50 mM phosphate buffer at pH 7.0, 8.0, and 9.0, as a function of weight fraction qP4VP.
  • Figure 13A depicts three components (i.e., a charged homopolymer, a quaternized P4VP homopolymer, and a protein) used to investigate coacervation.
  • Figure 13B depicts the results of DLS studies at 600 nm of the components from Figure 13A in 50 mM phosphate buffer at pH 7.0, 8.0, and 9.0, as a function of weight fraction qP4VP.
  • the 0.7 weight fraction was selected for DLS studies shown in Figure 14A and Figure 14B.
  • the 0.2 weight fraction was selected for DLS studies shown in Figure 15.
  • Figure 14A depicts three components (i.e., a charged homopolymer, a POEGMA-b- qP4VP block copolymer, and a protein) used to investigate micelle formation.
  • Figure 17A depicts DLS data of coacervate core micelle solutions (POEGMA-b- qP4VP mixed with PAA) that shows the percent mass of small scatterers (squares, likely complexes or free polymer), micelle-sized species (circles), and larger species (triangles, likely dust or aggregates) as a function of HEPES concentration, showing that the micelles are stable to conditions up to 50 mM HEPES.
  • POEGMA-b- qP4VP mixed with PAA shows the percent mass of small scatterers (squares, likely complexes or free polymer), micelle-sized species (circles), and larger species (triangles, likely dust or aggregates) as a function of HEPES concentration, showing that the micelles are stable to conditions up to 50 mM HEPES.
  • Figure 17B depicts DLS data of coacervate core micelle solutions (POEGMA-b- qP4VP mixed with PAA) at that shows the percent intensity of small scatterers (squares, likely complexes or free polymer), micelle-sized species (circles), and larger species (triangles, likely dust or aggregates) as a function of HEPES concentration, showing that the micelles are stable to conditions up to 50 mM HEPES.
  • POEGMA-b- qP4VP mixed with PAA shows the percent intensity of small scatterers (squares, likely complexes or free polymer), micelle-sized species (circles), and larger species (triangles, likely dust or aggregates) as a function of HEPES concentration, showing that the micelles are stable to conditions up to 50 mM HEPES.
  • Figure 18A depicts DLS data of coacervate core micelle solutions (POEGMA-b- qP4VP mixed with PAA) in 50 mM pH 8 HEPES buffer that shows the percent mass of small scatterers (squares), likely complexes or free polymer), micelle-sized species
  • Figure 18B depicts DLS data of coacervate core micelle solutions (POEGMA-b- qP4VP mixed with PAA) in 50 mM pH 8 HEPES buffer that shows the percent intensity of small scatterers (squares), likely complexes or free polymer), micelle-sized species
  • Figure 19 depicts small angle neutron scattering (SANS) of micelles with OPH protein in 50 mM pH 8 HEPES at total polymer concentration 20 mg/mL, OPH concentration at ⁇ 2 mg/mL.
  • SANS small angle neutron scattering
  • Figure 20 depicts the specific activities against paraoxon of OPH only, OPH with PAA, OPH with POEGMA-b-qP4VP, and OPH in micelles. This shows that OPH with the block copolymer and in the micelles retains its activity over time after treatment at 37 °C in
  • Figure 21A depicts DLS data showing percent mass of different species— Small scatterers (squares, free polymer or small micelles), micelle-sized species (circles), larger scatterers (point-down triangles, possibly cylindrical micelles), and large aggregates (point- up triangles) - as a function of HEPES concentration. There is little dependence of inverse micelle formation on salt concentration. Conditions: 4 mg/mL POEGMA-b-qP4VP in 4%
  • FIG. 21B depicts DLS data showing percent intensity of different species— Small scatterers (squares, free polymer or small micelles), micelle-sized species (circles), larger scatterers (point-down triangles, possibly cylindrical micelles), and large aggregates (point- up triangles) - as a function of HEPES concentration. There is little dependence of inverse micelle formation on salt concentration. Conditions: 4 mg/mL POEGMA-b-qP4VP in 4% HEPES buffer, 96% ethanol.
  • Figure 22A depicts DLS data showing percent mass of different species— Small scatterers (squares, free polymer or small micelles), micelle-sized species (circles), larger scatterers (point-down triangles, possibly cylindrical micelles), and large aggregates (point- up triangles) - as a function of HEPES concentration. There is little dependence of inverse micelle formation on salt concentration. Conditions: 4 mg/mL POEGMA-b-qP4VP in 4% HEPES buffer, 96% DMMP.
  • Figure 22B depicts DLS data showing percent intensity of different species— Small scatterers (squares, free polymer or small micelles), micelle-sized species (circles), larger scatterers (point-down triangles, possibly cylindrical micelles), and large aggregates (point- up triangles) - as a function of HEPES concentration. There is little dependence of inverse micelle formation on salt concentration. Conditions: 4 mg/mL POEGMA-b-qP4VP in 4% HEPES buffer, 96% DMMP.
  • Figure 23A depicts DLS data showing percent mass of different species— Small scatterers (squares, free polymer or small micelles), micelle-sized species (circles), larger scatterers (point-down triangles, possibly cylindrical micelles), and large aggregates (point- up triangles)— as a function of positive charge fraction.
  • Mostly precipitates form when the solution is made up of mostly PAA (due to its insolubility in organic solvents), that changes into a solution that is mostly micelles ( ⁇ 30 nm in radius) between f 0.2-0.6, and then into a mixture of micelles ( ⁇ 30 nm in radius) and larger structures (-200 nm in radius).
  • Figure 23B depicts DLS data showing percent intensity of different species— Small scatterers (squares, free polymer or small micelles), micelle-sized species (circles), larger scatterers (point-down triangles, possibly cylindrical micelles), and large aggregates (point- up triangles)— as a function of positive charge fraction.
  • Small scatterers squares, free polymer or small micelles
  • micelle-sized species circles
  • larger scatterers point-down triangles, possibly cylindrical micelles
  • large aggregates point- up triangles
  • Figure 24B depicts DLS data showing percent intensity of different species— Small scatterers (squares, free polymer or small micelles), micelle-sized species (circles), larger scatterers (point-down triangles, possibly cylindrical micelles), and large aggregates (point- up triangles)— as a function of positive charge fraction.
  • Small scatterers squares, free polymer or small micelles
  • micelle-sized species circles
  • larger scatterers point-down triangles, possibly cylindrical micelles
  • large aggregates point- up triangles
  • Figure 25 depicts the specific activity against paraoxon of OPH in 90% 50 mM pH 8 HEPES after treatment for 24 hours in 96% ethanol (left bar) and DMMP (right bar). These data shows that the block copolymer is able to stabilize against treatment with ethanol, but not DMMP. The micelles are able to stabilize against DMMP, which is a good simulant for organophosphate compounds.
  • Figure 26 has five panels (a-e) showing the supercharging of model proteins, (a) Model proteins selected, represented with electrostatic surface potential ( ⁇ 5 kT/e c ) at the solvent-accessible surface rendered from solutions of the linearized Poisson-Boltzmann equation using the Adaptive Poisson-Boltzmann Solver (APBS).
  • APBS Adaptive Poisson-Boltzmann Solver
  • Figure 27 has two panels (a and b) showing bulk coacervation with supercharged proteins, (a) Summary of the turbidity profiles as a funciton of charge fraction (for negatively charged proteins, circles) or polymer weight fraction (for positively charged proteins, triangles) in 10 mM tris buffer, pH 8.0 (a-Chymotrypsinogen (upper left), lysozyme (upper right), myoglobin (bottom left), RNase A (bottom right)), (b) Bright field optical micrographs showing the lack of phase separation, liquid coacervates, or solid precipitates resulting from mixing supercharged proteins with qPDMAEMA at the midpoint of bulk coacervation. Scale bars, 20 ⁇ .
  • Figure 28 has two panels (a and b) showing salt and pH titrations of RNase A coacervates.
  • (a) Turbidimetric pH titrations of RNase A with qP4VP at an ionic strength of 10 mM and protein-polymer ratio r 5.
  • Figure 29 has two panels (a and b) showing protein incorporation in the coacervate phase, (a) Protein partitioning in the coacervate phase as a function of the expected protein charge, (b) Incorporation of supercharged proteins in the coacervate phase as a function of the protein-to-polymer ratio.
  • Figure 30 has two panels (a and b) showing micelle formation as a funciton of charge fraction, (a) The percentage of micelles in solution as determined by DLS intensity is plotted as a funciton of charge fraction for (from left to right) chymotrypsinogen, lysozyme, myoglobin, and RNase A. (b) The average micelle radii are plotted as a function of charge fraction for the same proteins.
  • Figure 31 has three panels (a-c) showing the stability of the complex coacervate core micelles, (a) Thermal stability of the micelles was assayed by DLS. The percentage of micelles in solution (left) and average hydrodynamic radius (right) are plotten as a funciton of temperature, (b) The ability of the micelles to reform after lyophilization was confirmed by DLS measurements before and after lyophilizing RNase A C3Ms. (c) Stability of micelles with RNase A to increased ionic strength was investigated by DLS and the average R h is plotted as a function of NaCl concentration. DETAILED DESCRIPTION
  • the invention relates to compositions and methods for catalyzing the hydrolysis of organophosphates, such as G-series or V-series nerve agents. See Figure 1A and Figure IB.
  • the invention relates to a method of remediating bulk chemical warfare agents, for example, on-site remediation.
  • the invention relates to a composition, comprising a complex coacervate core micelle, which may act as a nanoreactor.
  • the ionic, hydrophilic core encapsulates enzymes and water, which are necessary for hydrolysis, while the neutral corona solubilizes the micelle in organic solvent.
  • a charged polymer is used, and may act to absorb acidic by-products (such as HF). See Figure 2.
  • Coacervation is a known phenomenon in colloid chemistry.
  • coacervation is the phenomenon of salting out or phase separation of lyophilic colloids into liquid droplets, rather than solid aggregates.
  • Coacervation of a polymeric ingredient can be brought about in a number of different ways, for example by a change in temperature, a change of pH, addition of a low molecular weight substance or addition of a second macromolecular substance.
  • Two types of coacervation have been defined: simple coacervation and complex coacervation. In general, simple coacervation deals with systems containing only one polymeric ingredient, while complex coacervation deals with systems containing more than one polymeric ingredient.
  • complex coacervation is the liquid-liquid phase separation that results when solutions of two oppositely charged macro-ions are mixed, resulting in the formation of a dense macro-ion-rich phase, the precursors of which are soluble complexes.
  • Variables such as temperature, pH, and concentration, may be used to induce polymer phase separation, so as to produce a suspension of complex coacervate micelles.
  • Active agents, dyes, or proteins, such as enzymes may be encapsulated in complex coacervate core micelles.
  • hydrophilic "core materials” e.g., enzymes
  • an aqueous nano- or micro-environment may be dispersed in organic solvents.
  • Hydrolases are enzymes that catalyze the hydrolysis of a chemical bond.
  • An example of a hydrolase is organophosphate hydrolase (also known as aryldialkylphosphatase (EC 3.1.8.1), organophosphor o us hydrolase, phosphotriesterase, and paraoxon hydrolase), which has a molecular weight of about 39.1 kDa, 7 lysine residues, or a pi of about 8.1, or a combination thereof.
  • organophosphate acid anhydrolase also known as organophosphor o us acid anhydrolase (OPAA)
  • the enzyme is found in a diverse range of organisms, including protozoa, squid and clams, mammals, and soil bacteria. A highly active form of the enzyme may be isolated from the marine bacteria Alteromonas undina.
  • Organophosphate acid anhydrolase has a molecular weight of about 50.8 kDa, 21 lysine residues, or a pi of about 6.1 , or a combination thereof.
  • Model enzymes which may mimic the size, shape, or surface characteristics of hydrolytic enzymes, are also described herein.
  • a-chymotrypsinogen may be used.
  • a-Chymotrypsinogen is a proteolytic enzyme and a precursor of chymotrypsin.
  • a-Chymotrypsinogen has a molecular weight of about 25.7 kDa, 14 lysine residues, or a pi of about 8.2, or a combination thereof.
  • a-amylase Another model enzyme is a-amylase.
  • An amylase is an enzyme that catalyzes the hydrolysis of starch into sugars.
  • a-Amylase has a molecular weight of about 47.0 kDa, 17 lysine residues, or a pi of about 5.6, or a combination thereof.
  • model enzymes include, but are not limited to, lysozyme and myoglobin.
  • Nerve agents are a class of phosphorus-containing organic chemicals (organophosphates) that disrupt the mechanism by which nerves transfer messages to organs. The disruption is caused by blocking acetylcholinesterase, an enzyme that normally destroys acetylcholine, a neurotransmitter.
  • organophosphates organic chemicals
  • Nerve agents can also be absorbed through the skin.
  • G-series and V-series There are two main classes of nerve agents: G-series and V-series.
  • G-series agents are non-persistent, and include GA (tabun), GB (sarin), GD (soman), and GF (cyclosarin). The structures of these G-series agent are shown below.
  • V-series agents are persistent, meaning that these agents do not degrade or wash away easily and can, therefore, remain on clothes and other surfaces for long periods. In use, this characteristic allows the V-agents to be used to blanket terrain to guide or curtail the movement of enemy ground forces. The consistency of these agents is similar to oil; as a result, the contact hazard for V-agents is primarily - but not exclusively - dermal.
  • Commonly known V-series agents are VE, VG, VM, VR, and VX, the structures of which are shown below.
  • butyrylcholinesterase When administered prophylactically, this enzyme stoichiometrically binds the nerve agent in the bloodstream before it can exert effects on the nervous system.
  • Decontamination technologies for safe disposal, facility and site cleanup, and destruction of stockpiles of organophosphate nerve agents are needed to protect the environment as well as the public.
  • the technology should have following properties: environmentally friendliness; capable of safe transportation, storage and handling, including long term stability; serve as a first response to protect the civilian population; be capable of restoring contaminated facilities; not affect the operation of sensitive electronic equipment; generate minimal toxic byproducts; and render treated materials suitable for disposal in a municipal waste stream.
  • the invention relates to a nanostructure, comprising, consisting essentially of, or consisting of:
  • the invention relates to any one of the nanostructures described herein, wherein the nanostructure comprises (i) a polyanionic polymer; a block copolymer; and an enzyme.
  • the invention relates to any one of the nanostructures described herein, wherein the nanostructure comprises (ii) a block copolymer; and a modified enzyme.
  • the invention relates to any one of the nanostructures described herein, wherein the nanostructure consists essentially of (i) a polyanionic polymer; a block copolymer; and an enzyme.
  • the invention relates to any one of the nanostructures described herein, wherein the nanostructure consists essentially of (ii) a block copolymer; and a modified enzyme.
  • the invention relates to any one of the nanostructures described herein, wherein the nanostructure consists of (i) a polyanionic polymer; a block copolymer; and an enzyme.
  • the invention relates to any one of the nanostructures described herein, wherein the nanostructure consists of (ii) a block copolymer; and a modified enzyme.
  • the invention relates to any one of the nanostructures described herein, wherein the nanostructure comprises (i) a polyanionic polymer; a block copolymer; an enzyme; and an aqueous liquid.
  • the invention relates to any one of the nanostructures described herein, wherein the nanostructure comprises (ii) a block copolymer; a modified enzyme; and an aqueous liquid. In certain embodiments, the invention relates to any one of the nanostructures described herein, wherein the nanostructure consists essentially of (i) a polyanionic polymer; a block copolymer; an enzyme; and an aqueous liquid.
  • the invention relates to any one of the nanostructures described herein, wherein the nanostructure consists essentially of (ii) a block copolymer; a modified enzyme; and an aqueous liquid.
  • the invention relates to any one of the nanostructures described herein, wherein the nanostructure consists of (i) a polyanionic polymer; a block copolymer; an enzyme; and an aqueous liquid.
  • the invention relates to any one of the nanostructures described herein, wherein the nanostructure consists of (ii) a block copolymer; a modified enzyme; and an aqueous liquid.
  • the invention relates to any one of the nanostructures described herein, wherein the block copolymer comprises a plurality of first repeat units, and a plurality of second repeat units;
  • the first repeat unit is ,
  • R is H, alkyl, halo, hydroxy, amino, nitro, or cyano
  • Y is alkyl
  • X ⁇ is an anion
  • R is H, alkyl, halo, hydroxy, amino, nitro, or cyano
  • p 2-20, inclusive.
  • the invention relates to any one of the nanostructures described herein, wherein p is 4-10, inclusive. In certain embodiments, the invention relates to any one of the nanostructures described herein, wherein p is 4, 5, 6, 7, 8, 9, or 10.
  • the invention relates to any one of the nanostructures described herein, wherein R is independently H or alkyl.
  • the invention relates to any one of the nanostructures described herein, wherein one instance of R is alkyl.
  • the invention relates to any one of the nanostructures described herein, wherein one instance of R is methyl, ethyl, n-propyl, or i-propyl.
  • the invention relates to any one of the nanostructures described herein, wherein one instance of R is methyl.
  • the invention relates to any one of the nanostructures described herein, wherein three instances of R are H.
  • the invention relates to any one of the nanostructures described herein, wherein R is H.
  • the invention relates to any one of the nanostructures described herein, wherein Y is Ci-Cio alkyl. In certain embodiments, the invention relates to any one of the nanostructures described herein, wherein Y is C 2 -C 6 alkyl. In certain embodiments, the invention relates to any one of the nanostructures described herein, wherein Y is n-butyl.
  • the invention relates to any one of the nanostructures described herein, wherein X ⁇ is halide, boron tetrafluoride, phosphorus tetrafluoride, phosphorus hexafluoride, alkylsulfonate, fluoroalkylsulfonate, arylsulfonate, bis(alkylsulfonyl)amide, bis(fluoroalkylsulfonyl)amide, bis(arylsulfonyl)amide, (fluoroalkylsulfonyl)(fluoroalkylcarbonyl)amide, nitrate, nitrite, sulfate, hydrogensulfate, alkyl sulfate, aryl sulfate, carbonate, bicarbonate, carboxylate, phosphate, hydrogen phosphate, dihydrogen phosphate, or hypochlorite.
  • the invention relates to any one of the nanostructures described herein, wherein X ⁇ is halide.
  • the invention relates to any one of the nanostructures described herein, wherein X ⁇ is bromide, chloride, or iodide.
  • the invention relates to any one of the nanostructures described herein, wherein X ⁇ is bromide. In certain embodiments, the invention relates to an one of the nanostructures
  • the molecular weight of the block is about 12 kDa to about 60 kDa.
  • the molecular weight of the block is about 15 kDa, about 20 kDa, about 25 kDa, about 30 kDa, about 35 kDa, about 40 kDa, about 45 kDa, about 50 kDa, or about 55 kDa.
  • the invention relates to any one of the nanostructures
  • the invention relates to an one of the nanostructures
  • the molecular weight of the block is about 6 kDa, about 8 kDa, about 10 kDa, about 12 kDa, about 14 kDa, about 16 kDa, about 18 kDa, about 20 kDa, about 22 kDa, about 24 kDa, about 26 kDa, or about 28 kDa.
  • the invention relates to any one of the nanostructures described herein, wherein the molecular weight of the block copolymer is about 18 kDa to about 74 kDa. In certain embodiments, the invention relates to any one of the nanostructures described herein, wherein the molecular weight of the block copolymer is about 20 kDa, about 25 kDa, about 30 kDa, about 35 kDa, about 40 kDa, about 45 kDa, about 50 kDa, about 55 kDa, about 60 kDa, about 65 kDa, or about 70 kDa.
  • the invention relates to any one of the nanostructures described herein, wherein the block copolymer further comprises a first end-group; and the
  • first end-group has the following structure:
  • the invention relates to any one of the nanostructures described herein, wherein the block copolymer further comprises a second end-group;
  • the second end-group has the following structure:
  • the invention relates to any one of the nanostructures described herein, wherein the block copolymer is a diblock copolymer.
  • the invention relates to any one of the nanostructures described herein wherein the block copolymer has the following structure:
  • the invention relates to any one of the nanostructures described herein, wherein n is about 60, about 70, about 80, about 90, about 100, about 110, about 120, about 130, or about 140.
  • the invention relates to any one of the nanostructures described herein, wherein m is about 70, about 80, about 90, about 100, about 110, about 120, about 130, about 140, about 150, about 160, about 170, about 180, or about 190.
  • the invention relates to any one of the nanostructures described herein, wherein the enzyme or modified enzyme is an organophosphate hydrolase or a modified organophosphate hydrolase. In certain embodiments, the invention relates to any one of the nanostructures described herein, wherein the enzyme or modified enzyme is an organophosphate acid anhydrolase or a modified organophosphate acid anhydrolase.
  • the invention relates to any one of the nanostructures described herein, wherein the enzyme or modified enzyme is a-chymotrypsinogen or modified a-chymotrypsinogen.
  • the invention relates to any one of the nanostructures described herein, wherein the enzyme or modified enzyme is an a-amylase or a modified a- amylase.
  • the invention relates to any one of the nanostructures described herein, wherein the enzyme or modified enzyme is a lysozyme or a modified lysozyme.
  • the invention relates to any one of the nanostructures described herein, the enzyme or modified enzyme is a myoglobin or a modified myoglobin.
  • the invention relates to any one of the nanostructures described herein, wherein the molecular weight of the enzyme or the modified enzyme is about 20 kDa to about 60 kDa.
  • the invention relates to any one of the nanostructures described herein, wherein the molecular weight of the enzyme or the modified enzyme is about 22 kDa, about 23 kDa, about 24 kDa, about 25 kDa, about 26 kDa, about 27 kDa, about 28 kDa, about 29 kDa, about 30 kDa, about 31 kDa, about 32 kDa, about 33 kDa, about 34 kDa, about 35 kDa, about 36 kDa, about 37 kDa, about 38 kDa, about 39 kDa, about 40 kDa, about 41 kDa, about 42 kDa, about 43 kDa, about 44 kDa, about 45 kDa, about 46 kDa, about 47 kDa, about 48 kDa, about 49 kDa, about 50 kDa, about 51 kDa, about 52 kDa, or about
  • the invention relates to any one of the nanostructures described herein, wherein the temperature of the nanostructure is about 18 °C to about 50 °C.
  • the invention relates to any one of the nanostructures described herein, wherein the temperature of the nanostructure is about 20 °C , about 22 °C, about 24 °C, about 26 °C, about 28 °C, about 30 °C, about 32 °C, about 34 °C, about 36 °C, about 38 °C, about 40 °C, about 42 °C, about 44 °C, about 46 °C, or about 48 °C. In certain embodiments, the invention relates to any one of the nanostructures described herein, wherein the temperature of the nanostructure is about 23 °C.
  • the invention relates to any one of the nanostructures described herein, wherein the temperature of the nanostructure is about 37 °C.
  • the invention relates to any one of the nanostructures described herein, wherein the positive charge fraction f is about 0.1 to about 0.5.
  • the invention relates to any one of the nanostructures described herein, wherein the positive charge fraction ⁇ is about 0.2 to about 0.4.
  • the invention relates to any one of the nanostructures described herein, wherein the positive charge fraction f is about 0.3.
  • the invention relates to any one of the nanostructures described herein, wherein the nanostructure is a nanostructure of form (i); and the polyanionic polymer is polyacrylic acid.
  • the invention relates to any one of the nanostructures described herein, wherein the molecular weight of the polyacrylic acid is about 2 kDa to about 10 kDa.
  • the invention relates to any one of the nanostructures described herein, wherein the molecular weight of the polyacrylic acid is about 3 kDa, about 4 kDa, about 5 kDa, about 6 kDa, about 7 kDa, about 8 kDa, or about 9 kDa.
  • the invention relates to any one of the nanostructures described herein, wherein the nanostructure is a nanostructure of form (ii); and the modified enzyme comprises at least one non-natural pendant anionic moiety.
  • the invention relates to any one of the nanostructures described herein, wherein the modified enzyme comprises at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least 10 non-natural pendant anionic moieties.
  • the invention relates to any one of the nanostructures described herein, wherein the modified enzyme comprises about two, about three, about four, about five, about six, about seven, about eight, about nine, about 10, about 11, about 12, about 13, about 14, about 15, about 16, about 17, about 18, about 19, or about 20 non- natural pendant anionic moieties.
  • the invention relates to any one of the nanostructures described herein, wherein the pendant anionic moiety is covalently bonded to a lysine residue.
  • the invention relates to any one of the nanostructures described herein, wherein the pendant anionic moiety is a carboxylate moiety.
  • the invention relates to any one of the nanostructures described herein, wherein the zeta potential of the modified enzyme is less than about -40 mV.
  • the invention relates to any one of the nanostructures described herein, wherein the zeta potential of the modified enzyme is from about -40 mV to about -70 mV.
  • the invention relates to any one of the nanostructures described herein, wherein the zeta potential of the modified enzyme is about -40 mV, about -50 mV, about -60 mV, or about -70 mV.
  • the invention relates to any one of the nanostructures described herein, wherein the nanostructure further comprises an aqueous liquid.
  • the invention relates to any one of the nanostructures described herein, wherein the aqueous liquid comprises a buffer.
  • the invention relates to any one of the nanostructures described herein, wherein the buffer is selected from the group consisting of: N-(2- acetamido)-2-aminoethanesulfonic acid (aces), N-(2-acetamido)iminodiacetic acid (ADA), acetate, 2-amino-2 -methyl- 1,3 -propanediol (AMPD), 2-amino-2-methyl-l-propanol (AMP), N,N-bis(2-hydroxyethyl)-2-aminoethanesulfonic acid (BES), 3-(N,N-bis[2- hydroxyethyl]amino)-2-hydroxypropanesulfonic acid (DIPSO), N,N-bis(2- hydroxyethyl)glycine (Bicine), bis(2-hydroxyethyl)iminotris(hydroxymethyl)methane (Bis- Tris), l,3-bis[tris(hydroxymethyl)methyl
  • the invention relates to any one of the nanostructures described herein, wherein the concentration of buffer in the aqueous liquid is about 10 mM to about 100 mM.
  • the invention relates to any one of the nanostructures described herein, wherein the concentration of buffer in the aqueous liquid is about 20 mM, about 30 mM, about 40 mM, about 50 mM, about 60 mM, about 70 mM, about 80 mM, or about 90 mM.
  • the invention relates to any one of the nanostructures described herein, wherein the concentration of buffer in the aqueous liquid is about 50 mM.
  • the invention relates to any one of the nanostructures described herein, wherein the pH of the aqueous liquid is about 6.5, about 7.0, about 7.5, about 8.0, about 8.5, about 9.0, or about 9.5.
  • the invention relates to any one of the nanostructures described herein, wherein the pH of the aqueous liquid is about 8.0.
  • the invention relates to any one of the nanostructures described herein, wherein the nanostructure is a nanostructure of form (i); and the enzyme is substantially encapsulated by the block copolymer.
  • the invention relates to any one of the nanostructures described herein, wherein the nanostructure is a nanostructure of form (i); and the polyanionic polymer is substantially encapsulated by the block copolymer.
  • the invention relates to any one of the nanostructures described herein, further comprising an aqueous liquid, wherein the nanostructure is a nanostructure of form (i); and the aqueous liquid is substantially encapsulated by the block copolymer.
  • the invention relates to any one of the nanostructures described herein, wherein the nanostructure is a nanostructure of form (ii); and the modified enzyme is substantially encapsulated by the block copolymer.
  • the invention relates to any one of the nanostructures described herein, further comprising an aqueous liquid, wherein the nanostructure is a nanostructure of form (ii); and the aqueous liquid is substantially encapsulated by the block copolymer.
  • the invention relates to any one of the nanostructures described herein, wherein the hydrodynamic radius of the nanostructure is about 1.5 nm to about 60 nm.
  • the invention relates to any one of the nanostructures described herein, wherein the hydrodynamic radius of the nanostructure is about 2 nm, about 3 nm, about 4 nm, about 5 nm, about 6 nm, about 7 nm, about 8 nm, about 9 nm, about 10 nm, about 11 nm, about 12 nm, about 13 nm, about 14 nm, about 15 nm, about 16 nm, about 17 nm, about 18 nm, about 19 nm, about 20 nm, about 21 nm, about 22 nm, about 23 nm, about 24 nm, about 25 nm, about 26 nm, about 27 nm, about 28 nm, about 29 nm, about 30 nm, about 31 nm, about 32 nm, about 33 nm, about 34 nm, about 35 nm, about 36 nm, about 37 nm, about 38 nm, about
  • the invention relates to any one of the nanostructures described herein, wherein the radius of the nanostructure, as determined by SANS, is about 10 nm to about 25 nm.
  • the invention relates to any one of the nanostructures described herein, wherein the radius of the nanostructure, as determined by SANS, is about 10 nm, about 11 nm, about 12 nm, about 13 nm, about 14 nm, about 15 nm, about 16 nm, about 17 nm, about 18 nm, about 19 nm, about 20 nm, about 21 nm, about 22 nm, about 23 nm, about 24 nm, or about 25 nm.
  • the invention relates to a composition, comprising, consisting essentially of, or consisting of: an organic phase, an aqueous liquid phase, and a plurality of any of the nanostructures described herein.
  • the invention relates to any of the compositions described herein, wherein the compositions comprises an organic phase, an aqueous liquid phase, and a plurality of any of the nanostructures described herein.
  • the invention relates to any of the compositions described herein, wherein the composition consists essentially of an organic phase, an aqueous liquid phase, and a plurality of any of the nanostructures described herein.
  • the invention relates to any of the compositions described herein, wherein the composition consists of an organic phase, an aqueous liquid phase, and a plurality of any of the nanostructures described herein.
  • the invention relates to any of the compositions described herein, wherein the organic phase is an organic liquid phase.
  • the invention relates to any of the compositions described herein, wherein the organic phase comprises ethanol.
  • the invention relates to any of the compositions described herein, wherein the organic phase consists essentially of ethanol.
  • the invention relates to any of the compositions described herein, wherein the organic phase consists of ethanol.
  • the invention relates to any of the compositions described herein, wherein the organic phase comprises DMMP.
  • the invention relates to any of the compositions described herein, wherein the organic phase consists essentially of DMMP.
  • the invention relates to any of the compositions described herein, wherein the organic phase consists of DMMP.
  • the invention relates to any of the compositions described herein, wherein the total concentration of block copolymer and polyanionic polymer in the composition is about 1 mg/mL to about 40 mg/mL.
  • the invention relates to any of the compositions described herein, wherein the total concentration of block copolymer and polyanionic polymer in the composition is about 2 mg/mL, about 4 mg/mL, about 6 mg/mL, about 8 mg/mL, about 10 mg/mL, about 12 mg/mL, about 14 mg/mL, about 16 mg/mL, or about 18 mg/mL. In certain embodiments, the invention relates to any of the compositions described herein, wherein the concentration of block copolymer in the composition is about 1 mg/mL to about 40 mg/mL.
  • the invention relates to any of the compositions described herein, wherein the concentration of block copolymer in the composition is about 2 mg/mL, about 4 mg/mL, about 6 mg/mL, about 8 mg/mL, about 10 mg/mL, about 12 mg/mL, about 14 mg/mL, about 16 mg/mL, or about 18 mg/mL.
  • the invention relates to any of the compositions described herein, wherein the concentration of enzyme or modified enzyme in the composition is about 1 mg/mL to about 40 mg/mL.
  • the invention relates to any of the compositions described herein, wherein the concentration of enzyme or modified enzyme in the composition is about 2 mg/mL, about 4 mg/mL, about 6 mg/mL, about 8 mg/mL, about 10 mg/mL, about 12 mg/mL, about 14 mg/mL, about 16 mg/mL, or about 18 mg/mL.
  • the invention relates to any of the compositions described herein, wherein the volume ratio of organic phase to aqueous liquid phase is about 99: 1 to about 90: 10.
  • the invention relates to any of the compositions described herein, wherein the volume ratio of organic phase to aqueous liquid phase is about 98:2: about 97:3, about 96:4, about 95:5, or about 94:6.
  • the invention relates to a method of hydrolyzing an organophosphorous compound, comprising contacting the organophosphorous compound with an effective amount of any of the nanostructures or compositions described herein.
  • the invention relates to a method of decontaminating an area or a device contaminated with an organophosphorous compound, comprising contacting the area or the device with an effective amount of any of the nanostructures or compositions described herein.
  • the invention relates to any of the methods described herein, wherein the area contaminated with the organophosphorus compound is on the skin of a human or an animal. In certain embodiments, the invention relates to any of the methods described herein, wherein the area contaminated with the organophosphorus compound is on the clothing of a human.
  • the invention relates to any of the methods described herein, wherein the method is a catalytic method.
  • the invention relates to any of the methods described herein, wherein the organophosphorus compound is selected from the group consisting of: GA, GB, GD, GF, VE, VG, VM, VR, and VX.
  • RAFT RAFT polymerization was used to synthesize homopolymer from 2-(dimethylamino)ethyl methacrylate (DMAEMA) (98%, Aldrich) with a narrow molecular weight distribution. DMAEMA was passed through a basic alumina column prior to polymerization to remove inhibitors. 2-hydroxy ethyl 2-(((ethylthio)carbonothioyl)thio)-2-methylpropanoate (EMP- OH) was prepared as the RAFT chain transfer agent as described in the Supporting Information.
  • DMAEMA 2-(dimethylamino)ethyl methacrylate
  • EMP- OH 2-hydroxy ethyl 2-(((ethylthio)carbonothioyl)thio)-2-methylpropanoate
  • PDMAEMA 8 g was quaternized with iodomethane (99%, Sigma- Aldrich, 13 mL) in N,N-dimethylformamide (DMF). The reaction mixture was stirred at room temperature for 24 h and then the modified polymer was precipitated in diethyl ether and dried under vacuum. The degree of quaternization was >95% as determined by 1H NMR.
  • 4VP 4-vinylpyridine
  • OEGMA, M n 300 g/mol
  • Aldrich oligo(ethylene glycol) methyl ether methacrylate
  • the polymerization was carried out in a sealed flask at 75 °C and terminated after 6 h by removal of heat and exposure to oxygen. The polymer was then precipitated in diethyl ether and dried under vacuum.
  • the block copolymer was quaternized with iodomethane in DMF. The reaction mixture was stirred at room temperature for 24 h and then the modified polymer was precipitated in diethyl ether and dried under vacuum. The degree of quaternization was > 95% as determined by 1H NMR.
  • Sample preparation Stock solutions of qPDMAEMAi 3 3, qP4VPn 4 , and POEGMAi24-b-qP4VPio8 were prepared by dissolving the polymers in 10 mM tris buffer, pH 8.0. Protein solutions were prepared by diluting the initial solution of protein in 10 mM tris buffer, pH 8.0 to the desired final concentration. All solutions were filtered through 0.20 ⁇ inorganic membrane syringe filters (Whatman).
  • Turbidimetric titrations Protein and polymer samples were prepared at 2 mg/mL in lO mM tris buffer, pH 8.0. Turbidity was used to qualitatively measure the extent of complex formation as a function of charge stoichiometry and salt concentration. To vary the charge stoichiometry, the protein and polymer solutions were mixed at ratios varying from 100% protein to 100% polymer in 4% increments of polymer. Samples were prepared in triplicate in a 96 well plate and the percent transmittance was measured on a plate reader at 600 nm or 750 nm (myoglobin). Error bars on the plots represent the calculated standard deviation of the data.
  • Protein and polymer samples were prepared at 2 mg/mL in 10 mM tris buffer, pH 8.0. The samples were mixed at the protein-to-polymer ratio in the middle of the coacervation range determined by turbidimetric titration. If the protein sample did not form a coacervate phase it was mixed at the ratio determined for the protein closest in charge that did coacervate. The protein and polymer samples were briefly vortexed and then centrifuged for 10 minutes at 14,800 rpm. The dilute phase was removed by pipet and the coacervate phase was resuspended in an equal volume of 10 mM tris buffer, pH 8.0 with 500 mM NaCl.
  • the protein concentration was determined in triplicate in both the dilute and coacervate phases using a BCA assay (Pierce) following the manufacturer's instructions. Dynamic light scattering measurements. Protein and polymer samples were prepared at 4 mg/mL in 10 mM tris buffer, pH 8.0. Encapsulation of proteins with the block copolymer was achieved by first diluting the protein stock solution in 10 mM tris buffer, pH 8.0 to the desired concentration, followed by addition of the polymer. After mixing, samples were allowed to equilibrate at 4 °C for at least 24 h before measurements. For each sample, DLS measurements were repeated three times and involved collection of 5-10 light scattering intensity fluctuation traces. Additionally, samples were prepared in triplicate. Error bars on the plots represent the calculated standard deviation of the data. For micellar compositions, when the standard deviation in the average radius was larger than 3 nm, the value was determined to be unreliable and the radius value was not plotted.
  • Example 2 Improving coacervation by adding a polyanionic polymer
  • 2,2'-Azobisisobutyronitrile (AIBN, Aldrich, 98%>) was purified by recrystallization from ethanol.
  • 01igo(ethylene glycol) methyl ether methacrylate (OEGMA, average Mn 300 g/mol, Aldrich) and 4-vinyl pyridine (4 VP, Aldrich, > 95%>) were purified over basic alumina prior to use.
  • OPH expression and Purification OPH gene in pET15b expression plasmid (Novagen, USA) was expressed and purified as described elsewhere with minor modifications. OPH enzymes were expressed in the presence of 1 mM CoCl 2 only for the last 3 hours of expression. Resuspension of the frozen cells was done without DNase or RNase A, and 0.25 mg/mL lysozyme was added instead. OPH enzymes were dialyzed into 50 mM HEPES (pH 8), and 0.1 mM CoCl 2 .
  • Block copolymers were synthesized in a two- step reversible addition-fragmentation chain transfer (RAFT) polymerization, followed by a quaternization process.
  • RAFT reversible addition-fragmentation chain transfer
  • CPP 4-cyano-4- (phenylcarbonothioylthio)pentanoic acid
  • Aqueous polymer-only micelle samples were mixed at room temperature and then mixed at 4 °C overnight prior to measurement. Samples were allowed to come to room temperature prior to measurement. Aqueous micelle samples containing protein were prepared by first mixing PAA and the protein of interest for at least 30 minutes. Block copolymer was then added, and the sample was allowed to mix overnight at 4 °C. All protein samples were kept at 4 °C until just before measurements were performed. All samples containing organic solvents were prepared at 100 mg/mL (25X) polymer in pH 8 HEPES buffer of specified concentration and allowed to equilibrate overnight at this concentration. Organic solvent content was added in such that the increase in organic solvent content of the solution never exceeded 10%.
  • All samples were prepared at a total polymer concentration of 4 mg/mL unless otherwise specified. All aqueous polymer and protein samples were filtered through a 0.1 ⁇ syringe filter prior to mixing to prevent dust from affecting the integrity of light scattering data. All DMMP and ethanol solvent used were filtered through a 0.45 ⁇ and 0.1 ⁇ filter prior to addition to aqueous samples. All resuspended lyophilized samples were refiltered through a 0.45 ⁇ filter prior to measurement.
  • Small-Angle Neutron Scattering Small-angle neutron scattering (SANS) samples were prepared in deuterated water and ethanol at 20 mg/mL total polymer concentration and 2 mg/mL total OPH concentration.
  • the dilution factor refers to the amount of dilution performed on the OPH solution
  • C is the concentration of OPH in the original solution (mg/L)
  • 1.0 is the path length of the light (cm)
  • 17100 is the molar extinction coefficient of /?-nitrophenol (M 1 cm "1 ).

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Abstract

L'invention concerne des micelles à noyau de coacervat complexe comprenant une enzyme qui peut hydrolyser des composés organophosphorés, tels que des agents neurotoxiques, et, par exemple, leur utilisation dans la remédiation ou la décontamination de réserves d'armes chimiques.
PCT/US2015/050387 2014-09-16 2015-09-16 Micelles à noyau de coacervat complexe à copolymère bloc pour la catalyse enzymatique dans un solvant organique WO2016044400A1 (fr)

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CN109988780A (zh) * 2019-03-19 2019-07-09 苏州大学 基于甲基丙烯酸缩水甘油酯的高性能基因载体及其应用

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